Registration Dossier
Registration Dossier
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EC number: 477-690-9 | CAS number: 874819-71-3
Endpoint summary
Administrative data
Key value for chemical safety assessment
Genetic toxicity in vitro
Description of key information
Ames-Test: positiv in Salmonella TyphimuriumTA 1537 (without metabolic activation) and TA 98 (with and without metabolic activation).
Mouse lymphoma test: negative
In vivo micronucleus test: negative
Link to relevant study records
- Endpoint:
- in vitro cytogenicity / micronucleus study
- Data waiving:
- study scientifically not necessary / other information available
- Justification for data waiving:
- an in vitro cytogenicity study in mammalian cells or in vitro micronucleus study does not need to be conducted because adequate data from an in vivo cytogenicity test are available
- Endpoint:
- in vitro gene mutation study in bacteria
- Remarks:
- Type of genotoxicity: gene mutation
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 2004-08-24 - 2004-10-25
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Reason / purpose for cross-reference:
- reference to other study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 471 (Bacterial Reverse Mutation Assay)
- Version / remarks:
- OECD 471 (1997)
- GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- bacterial reverse mutation assay
- Species / strain / cell type:
- S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
- Species / strain / cell type:
- E. coli WP2 uvr A
- Metabolic activation:
- with and without
- Metabolic activation system:
- S9 liver microsomal fraction
- Test concentrations with justification for top dose:
- Concentration range in the main test (with metabolic activation): 50 ... 5000 µg/plate
Concentration range in the main test (without metabolic activation): 50 ... 5000 µg/plate - Vehicle / solvent:
- Solvent: dimethyl sulfoxide
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- yes
- Positive controls:
- yes
- Positive control substance:
- sodium azide
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- yes
- Positive controls:
- yes
- Positive control substance:
- 9-aminoacridine
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- yes
- Positive controls:
- yes
- Positive control substance:
- 2-nitrofluorene
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- yes
- Positive controls:
- yes
- Positive control substance:
- methylmethanesulfonate
- Key result
- Species / strain:
- S. typhimurium TA 1535
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Key result
- Species / strain:
- S. typhimurium TA 1537
- Metabolic activation:
- without
- Genotoxicity:
- positive
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Key result
- Species / strain:
- S. typhimurium TA 1537
- Metabolic activation:
- with
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Key result
- Species / strain:
- S. typhimurium TA 98
- Metabolic activation:
- with and without
- Genotoxicity:
- positive
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Species / strain:
- S. typhimurium TA 100
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- not valid
- Positive controls validity:
- not valid
- Species / strain:
- E. coli WP2 uvr A
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Additional information on results:
- Observations:
An increase in revertant numbers was observed at concentrations from 5000 to 500 µg/plate of the test item
in the bacterial strain Salmonella typhimurium TA 1537
in the experiments without metabolic activation as well
as in the plate incorporation test and in the preincubation
test and in the bacterial strain Salmonella typhimurium
TA 98 in all test variants.
This increase was confirmed by a repetition of all positive
experiments. - Remarks on result:
- other: all strains/cell types tested
- Conclusions:
- Interpretation of results :
positive
N-(2-Nitrophenyl)phosphoric triamide induces gene mutation in Salmonella Typhimurium TA 1537 and TA 98 under the experimental conditions described. - Executive summary:
N-(2-Nitrophenyl)phosphoric triamide was tested for a possible potential to induce gene mutation in bacteria according to OECD guideline 471.
As test organisms theSalmonella typhimuriumstrains TA 98, TA 100, TA 1535, TA 1537 and theEscherichia colistrain WP2uvrA were used. The bacteria were exposed toN-(2-Nitrophenyl)phosphoric triamideboth, in the presence and absence of rat liver S9 metabolic activation. Two independent experiments were performed according to the standard plate incorporation method (experiment I) or the pre-incubation method (experiment II), respectively.
The concentrations 5000, 1000, 500, 100 and 50 µg/plate were chosen for the experiments according to the standard plate incorporation method and for the experiments according to the pre-incubation method.
Not any cytotoxicity or precipitation was observed in all experiments.
An untreated, a vehicle control and appropriate positive controls were included into the experimental design. Triplicate plates were scored for each experimental point.
The revertant frequencies of the negative controls were within the expected range and the positive control chemicals induced marked increases in revertant colonies.
N-(2-Nitrophenyl)phosphoric triamideinduced at some experimental points (from 5000 to 500 µg/plate) a reproducible, dose dependent, statistically significant increase of revertant numbers for the bacterial strainSalmonella typhimuriumTA 1537 in the experiments without metabolic activation as well as in the plate incorporation test and in the pre-incubation test and for the bacterial strainSalmonella typhimuriumTA 98 in the experiments with and without metabolic activation as well as in the plate incorporation test and in the pre-incubation test.
Based on the results of the reported study it is concluded that N-(2-Nitrophenyl)phosphoric
triamide induces gene mutation inSalmonella typhimurium under the experimental conditions described.
- Endpoint:
- in vitro gene mutation study in mammalian cells
- Remarks:
- Type of genotoxicity: gene mutation
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 2009-01-23 - 2009-05-18
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: The GLP study was conducted according to an internationally accepted guideline. All study parameters are based on the specific guideline.
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- mammalian cell gene mutation assay
- Target gene:
- autosomal thymidine kinase (TK) locus of heterozygous L5178Y/TK+/" cells
- Species / strain / cell type:
- mouse lymphoma L5178Y cells
- Metabolic activation:
- with and without
- Metabolic activation system:
- Phenobarbital/p-Naphthoflavone induced rat liver S9
- Test concentrations with justification for top dose:
- Experiment 1:
without S9: 52.5, 105, 210, 420, 840, 1680 µg/l
with S9: 52.5, 105, 210, 420, 840, 1680 µg/l
Experiment 2:
without S9: 105, 210, 420, 840, 1680, 2200 µg/l
with S9: 52.5, 105, 210, 420, 840, 1680 µg/l - Vehicle / solvent:
- DMSO
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- yes
- Positive controls:
- yes
- Positive control substance:
- cyclophosphamide
- methylmethanesulfonate
- Details on test system and experimental conditions:
- Prior to mutagenicity testing the amount of spontaneous mutants is reduced by growing the cells for one day in RPMI 1640-HAT medium supplemented with:
hypoxanthine 1.0x10-4M
aminopterin 2.0x10-7M
thymidine 1.6x10-5M
The incubation of the cells in HAT-medium is followed by a recovery period of 2 days in RPMI 1640 medium containing:
hypoxanthine 1.0x10-4M
thymidine 1.6x10-5M
After this incubation the L5178Y cells are returned to normal RPMI 1640 medium (complete culture medium).
Large stocks of the cleansed L5178Y cell line are stored in liquid nitrogen in the cell bank of Harlan CCR allowing the repeated use of the same cell culture batch in many experiments. Before freezing, each batch was screened for mycoplasma contamination and checked for karyotype stability. Consequently, the parameters of the experiments remain similar because of the reproducible characteristics of the cells.
Thawed stock cultures are propagated in plastic flasks in RPMI 1640 complete culture medium. The cells are subcultured two times prior to treatment. The cell cultures are incubated at 37 ± 1.5°C in a humidified atmosphere with 4.5 % carbon dioxide and 95.5 % ambient air. - Evaluation criteria:
- A test item is classified as mutagenic if the induced mutation frequency reproducibly exceeds a threshold of 126 colonies per 106 cells above the corresponding solvent control or negative control, respectively.
A relevant increase of the mutation frequency should be dose-dependent.
A mutagenic response is considered to be reproducible if it occurs in both parallel cultures.
However, in the evaluation of the test results the historical variability of the mutation rates in negative and vehicle controls and the mutation rates of all negative and vehicle controls of this study are taken into consideration.
Results of test groups are generally rejected if the relative total growth, and the cloning efficiency 1 is less than 10 % of the vehicle control unless the exception criteria specified by the IWGT recommendations are fulfilled.
Whenever a test item is considered mutagenic according to the above mentioned criteria, the ratio of small versus large colonies is used to differentiate point mutations from clastogenic effects. If the increase of the mutation frequency is accompanied by a reproducible and dose dependent shift in the ratio of small versus large colonies clastogenic effects are indicated. - Statistics:
- A linear regression (least squares) was performed to assess a possible dose dependent increase of mutant frequencies using SYSTAT®11 (SYSTAT Software, Inc., 501, Canal Boulevard, Suite C, Richmond, CA 94804, USA) statistics software. The number of mutant colonies obtained for the groups treated with the test item was compared to the solvent control groups. A trend is judged as significant whenever the p-value (probability value) is below 0.05. However, both, biological relevance and statistical significance were considered together.
- Key result
- Species / strain:
- mouse lymphoma L5178Y cells
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- other: all strains/cell types tested
- Remarks:
- Migrated from field 'Test system'.
- Conclusions:
- Interpretation of results (migrated information):
negative
During the mutagenicity test described and under the experimental conditions reported the test item did not induce mutations in the mouse lymphoma thymidine kinase locus assay using the cell line L5178Y in the absence and presence of metabolic activation.
Therefore, N-(2-nitrophenyl)phosphoricacid triamide (P 101/04) is considered to be non-mutagenic in this mouse lymphoma assay. - Executive summary:
The study according to OECD guideline 476 was performed to investigate the potential of N-(2-nitrophenyl)phosphoricacid triamide (P 101/04) to induce mutations at the mouse lymphoma thymidine kinase locus using the cell line L5178Y. The assay was performed in two independent experiments, using two parallel cultures each. The first main experiment was performed with and without liver microsomal activation and a treatment period of 4 h. The second experiment was performed with a treatment period of 24 hours in the absence of metabolic activation and 4 hours in the presence of metabolic activation. The highest applied concentration (1680 (jg/mL) was chosen with regard to the solubility properties of the test item in organic solvents and aqueous media. Precipitation or turbidity of the test item was noted in all experimental parts at the maximum concentration. No substantial and reproducible dose dependent increase in mutant colony numbers was observed in both main experiments. No relevant shift of the ratio of small versus large colonies was observed up to the maximal concentration of the test item. Appropriate reference mutagens were used as positive controls and showed a distinct increase in induced mutant colonies, indicating that the tests were sensitive and valid.
N-(2-nitrophenyl)phosphoricacid triamide (P 101/04) is considered to be non-mutagenic in this mouse lymphoma assay.
Referenceopen allclose all
Bacteria tester strain |
MA |
Exp. |
Mean numb |
ers of revertants per plate |
Evaluation |
||||||
Controls |
µg test item per plate |
||||||||||
UTC |
VC |
PC |
5000 |
1000 |
500 |
100 |
50 |
||||
Salmonella typhimurium |
|||||||||||
TA1535 |
no |
1St |
17.3 |
16.0 |
1616.0 |
14.3 |
15.7 |
14.3 |
17.7 |
19.3 |
- |
no |
2nd |
8.0 |
7.3 |
710.0 |
8.0 |
10.7 |
11.3 |
11.0 |
8.0 |
- |
|
yes |
1st |
21.0 |
13.0 |
162.7 |
15.0 |
17.0 |
16.3 |
12.3 |
15.0 |
- |
|
yes |
2nd |
11.3 |
11.7 |
156.3 |
8.0 |
11.0 |
13.7 |
9.0 |
10.0 |
- |
|
TA1537 |
no |
1st |
7.7 |
7.3 |
1973.3 |
15.7 |
9.7 |
13.0 |
10.7 |
6.0 |
+ |
no |
1str |
16.3 |
20.7 |
2368.0 |
34.0 |
18.3 |
n.e. |
n.e. |
n.e. |
+ |
|
no |
2nd |
12.3 |
11.3 |
2789.3 |
72.3 |
23.7 |
21.0 |
17.0 |
12.7 |
+ |
|
no |
2r |
11.3 |
13.7 |
1170.7 |
72.3 |
26.3 |
n.e. |
n.e. |
n.e. |
+ |
|
yes |
1st |
14.7 |
9.0 |
478.0 |
16.3 |
9.7 |
10.3 |
11.7 |
11.7 |
- |
|
yes |
2nd |
10.3 |
9.7 |
384.0 |
14.3 |
7.7 |
12.7 |
7.0 |
6.3 |
- |
|
TA98 |
no |
2St |
16.7 |
19.7 |
473.7 |
80.7 |
40.3 |
27.0 |
20.3 |
20.0 |
+ |
no |
1st1r |
18.3 |
20.3 |
566.7 |
71.0 |
36.3 |
26.3 |
n.e. |
n.e. |
+ |
|
no |
2"d |
21.7 |
20.3 |
476.0 |
176.0 |
55.0 |
33.0 |
27.3 |
31.3 |
+ |
|
no |
2ndr |
17.3 |
9.7 |
504.0 |
166.0 |
53.7 |
19.3 |
n.e. |
n.e. |
+ |
|
vcs |
1st |
27.7 |
27.0 |
2245.3 |
66.7 |
38.7 |
26.7 |
29.7 |
22.0 |
t- |
|
ves |
1str |
31.7 |
23.7 |
2245.3 |
75.7 |
28.7 |
27.3 |
32.7 |
n.e. |
+ |
|
yes |
2nd |
12.7 |
11.3 |
2688.0 |
29.0 |
18.0 |
13.0 |
12.3 |
11.3 |
+ |
|
yes |
2ndr |
33.0 |
29.0 |
2634.7 |
52.3 |
41.7 |
29.0 |
22.7 |
n.e. |
+ |
|
TA100 |
no |
1st |
192.0 |
192.3 |
1701.3 |
191.3 |
174.3 |
192.7 |
188.0 |
183.3 |
- |
no |
2nd |
129.3 |
111.0 |
1552.0 |
152.7 |
111.3 |
125.3 |
120.0 |
128.3 |
- |
|
yes |
1st |
168.7 |
149.3 |
1365.3 |
188.0 |
164.7 |
184.7 |
158.3 |
150.0 |
- |
|
yes |
2nd |
130.3 |
122.3 |
2613.3 |
136.7 |
122.7 |
133.3 |
120.0 |
116.3 |
- |
|
Escherichia coli |
|||||||||||
WP2 uvrA |
no |
1st |
29.0 |
21.0 |
304.3 |
17.3 |
19.7 |
27.7 |
29.7 |
26.3 |
- |
no |
2nd |
30.0 |
33.3 |
521.3 |
21.7 |
26.0 |
26.3 |
24.7 |
35.0 |
- |
|
yes |
1st |
32.3 |
35.3 |
131.3 |
28.3 |
30.0 |
28.3 |
31.3 |
28.0 |
- |
|
yes |
2nd |
28.0 |
32.7 |
155.3 |
30.5 |
31.3 |
31.7 |
40.7 |
36.0 |
- |
|
|
|
|
relative |
mutant |
|
relative |
mutant |
|
|
conc. pg |
S9 |
total |
colonies/ |
|
total |
colonies/ |
|
|
per mL |
mix |
growth |
106cells |
threshold |
growth |
106cells |
threshold |
Column |
1 |
2 |
3 |
4 |
5 |
6 |
7 |
8 |
Experiment I/ 4htreatment |
|
culture I |
culture II |
|||||
Solv. control with DMSO |
|
- |
100.0 |
134 |
260 |
100.0 |
151 |
277 |
Pos. control with MMS |
13.0 |
- |
36.3 |
297 |
260 |
35.4 |
358 |
277 |
Test item |
52.5 |
- |
culture was not continued* |
culture was not continued* |
||||
Test item |
105.0 |
- |
97.6 |
83 |
260 |
84.1 |
191 |
277 |
Test item |
210.0 |
- |
110.0 |
115 |
260 |
77.4 |
274 |
277 |
Test item |
420.0 |
- |
91.8 |
105 |
260 |
121.8 |
233 |
277 |
Test item |
840.0 |
- |
60.6 |
121 |
260 |
84.4 |
204 |
277 |
Test item |
1680(p) |
- |
113.7 |
78 |
260 |
67.2 |
175 |
277 |
|
|
|
|
|
|
|
|
|
Solv. control with DMSO |
|
+ |
100.0 |
110 |
236 |
100.0 |
84 |
210 |
Pos. control with CPA |
3.0 |
+ |
32.1 |
307 |
236 |
26.0 |
184 |
210 |
Pos. control with CPA |
4.5 |
+ |
6.9 |
276 |
236 |
23.5 |
291 |
210 |
Test item |
52.5 |
+ |
culture was not continued* |
culture was not continued* |
||||
Test item105.0 |
+ |
107.9 |
124 |
236 |
62.0 |
91 |
210 |
|
Test item |
210.0 |
+ |
104.0 |
117 |
236 |
69.7 |
115 |
210 |
Test item |
420.0 |
+ |
83.9 |
145 |
236 |
67.5 |
127 |
210 |
Test item |
840.0 |
+ |
91.6 |
114 |
236 |
46.0 |
120 |
210 |
Test item |
1680(p) |
+ |
72.6 |
111 |
236 |
59.0 |
OA U\J |
210 |
Experiment11 / 24htreatment |
|
culture I |
culture II |
|||||
Solv. control with DMSO |
|
- |
100.0 |
109 |
235 |
100.0 |
224 |
350 |
Pos. control with MMS |
13.0 |
- |
22.4 |
459 |
235 |
44.2 |
448 |
350 |
Test item |
105.0 |
- |
88.1 |
150 |
235 |
126.5 |
152 |
350 |
Test item |
210.0 |
- |
59.1 |
199 |
235 |
91.0 |
183 |
350 |
Test item |
420.0 |
- |
70.7 |
118 |
235 |
108.6 |
122 |
350 |
Test item |
840.0 |
- |
35.3 |
161 |
235 |
48.3 |
183 |
350 |
Test item |
1680.0 |
- |
5.7 |
321 |
235 |
18.4 |
178 |
350 |
Test item |
2200(p) |
- |
culture was not continued** |
culture was not continued* |
||||
Experiment II/ 4htreatment |
|
culture I |
culture II |
|||||
Solv. control with DMSO |
|
+ |
100.0 |
88 |
214 |
100.0 |
92 |
218 |
Pos. control with CPA |
3.0 |
+ |
53.6 |
247 |
214 |
36.7 |
158 |
218 |
Pos. control with CPA |
4.5 |
+ |
24.7 |
314 |
214 |
18.3 |
308 |
218 |
Test item |
52.5 |
+ |
culture was not continued* |
culture was not continued* |
||||
Test item |
105.0 |
+ |
192.2 |
86 |
214 |
89.7 |
73 |
218 |
Test item |
210.0 |
+ |
153.4 |
78 |
214 |
82.7 |
49 |
218 |
Test item |
420.0 |
+ |
129.3 |
132 |
214 |
54.4 |
74 |
218 |
Test item |
840.0 |
+ |
132.6 |
106 |
214 |
56.6 |
65 |
218 |
Test item |
1680(p) |
+ |
96.9 |
121 |
214 |
73.1 |
48 |
218 |
Endpoint conclusion
- Endpoint conclusion:
- adverse effect observed (positive)
Genetic toxicity in vivo
Description of key information
Ames-Test: positiv in Salmonella TyphimuriumTA 1537 (without metabolic activation) and TA 98 (with and without metabolic activation).
Mouse lymphoma test: negative
In vivo micronucleus test: negative
Link to relevant study records
- Endpoint:
- in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
- Remarks:
- Type of genotoxicity: gene mutation
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 2004-11-29 - 2005-02-01
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.12 (Mutagenicity - In Vivo Mammalian Erythrocyte Micronucleus Test)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- micronucleus assay
- Species:
- mouse
- Strain:
- NMRI
- Sex:
- male/female
- Details on test animals or test system and environmental conditions:
- Species / Strain: Mouse, NMRI
Sex:male, female (nulliparous, non-pregnant)
Supplier:Charles River Wiga GmbH, D-97320 Sulzfeld
Hygiene status upon supply: SPF
Age at start of acclimatisation: 6-8 weeks
Acclimatisation:7 days before start of dosing (pre-experiment for toxicity) 14 days before start of dosing (main experiment)
Mean body weight on day of administration:Males:31.4 g± 1.7g(5.4%) n = 25; Females: 27.5 g± 1.4 g (5.3%) n=25 - Route of administration:
- oral: gavage
- Vehicle:
- 0.5 % (m/v) solution of Tylose MH 1000 in deionised water
- Details on exposure:
- oral administration using a metal catheter, single dose, 10 ml/kg body weight
- Duration of treatment / exposure:
- single dose
- Frequency of treatment:
- single dose
- Remarks:
- Doses / Concentrations:
2000 mg/kg bw
Basis:
actual ingested - No. of animals per sex per dose:
- 5 males and 5 females
- Positive control(s):
- Cyclophosphamide (CPA)
- Tissues and cell types examined:
- Bone Marrow Smears
- Details of tissue and slide preparation:
- Preparation of Bone Marrow Smears
At the appropriate termination time, the animals were killed by cervical dislocation.
Bone marrow was removed from the femurs using 1 ml of fetal calf serum, each, sedimented using centrifugation at 1200 Rpm for 10 minutes (approximately 700 g) and smears prepared on microscope slides. The smears were aged for approximately 24 hours before staining with May-Grunwald/Giemsa solution. After this staining the polychromatic erythrocytes (PCE) appear bluish, normochromatic erythrocytes (NCE) appear pink to yellowish and micronuclei appears reddish-violet. - Evaluation criteria:
- All slides were coded before scoring and scored blind. A minimum of 2000 polychromatic erythrocytes (PCE) was scored for the presence of micronuclei (= MPCE = micronucleated polychromatic erythrocytes) for each animal. The proportion of PCEs among total erythrocytes (PCEs + normochromatic erythrocytes [NCE]) was determined for each animal on the basis of 200 erythrocytes.
The test item is classified mutagenic, if it induces a statistically significant increase at the sampling times of 24 or 48 hours with biological relevance (>0.4 % micronuclei per animal per dose group). A statistically significant increase might require further confirmation by the demonstration of a dose response relationship at the respective sampling time. - Statistics:
- Micronucleus scores (MCPE) and the proportion of PCEs among total erythrocytes are presented as individual values. In addition group means and standard deviations are calculated for each sex and experimental group.
The statistical significance compared to the vehicle control were proved by means the Welch t-test (Rasch et al., Verfahrensbibliothek Versuchsplanung und -auswertung, Berlin 1981).
Statistical significance is declared at the 5 % level (one-sided). - Key result
- Sex:
- male/female
- Genotoxicity:
- negative
- Toxicity:
- no effects
- Vehicle controls validity:
- valid
- Positive controls validity:
- valid
- Conclusions:
- Interpretation of results : negative
Based on the results of the reported study, it is concluded that N-(2-nitrophenyl)phosphoric triamide does not induce an increase of micronuclei in polychromatic erythrocytes of NMRI-mice under the experimental conditions described.
P 101/04 is therefore considered to be non-mutagenic in the mouse bone marrow micronucleus test. - Executive summary:
In a mouse micronucleus test following EC-guideline B.12 under GLP conditions, the ability of N-(2-Nitrophenyl)phosphoric triamide to induce micronuclei was investigated.
Groups of 5 male and 5 female NMRI-mice were exposed to P 101/04 at the limit-test dose of 2000 mg/kg body weight. The test item was administered orally using a metal catheter. A 0.5 % (m/v) solution of Tylose MH 1000 in deionised water (10 ml/kg body weight) served as vehicle control and Cyclophosphamide (CPA) at a dose of 40 mg/kg body weight - also administered orally - was used as positive control. Bone marrow smears were prepared at 24 hours (vehicle control, positive control, dose group) and at 48 hours (vehicle control, dose group) after dosing. Two thousand polychromatic erythrocytes per animal were analysed for the presence of micronuclei. To investigate bone marrow toxicity the proportion of polychromatic erythrocytes among total erythrocytes was evaluated on the basis of 200 erythrocytes. The frequency of micronucleated polychromatic erythrocytes (MPCEs) in the vehicle control group was within the physiological range. Treatment with CPA induced statistically significant increases in the incidence of MPCEs. In none of the experimental groups treated with N-(2-nitrophenyl)phosphoric triamide
an increase in MPCEs or a statistically significant change of the proportion of polychromatic erythrocytes among total erythrocytes was observed. Based on the results of the study reported it is concluded that
N-(2-nitrophenyl)phosphoric triamide
does not induce micronuclei in polychromatic erythrocytes of NMRI-mice under the described experimental conditions.
N-(2-nitrophenyl)phosphoric triamidei
s therefore considered to be non-mutagcnic in the mouse bone marrow micronucleus test.
Reference
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed (negative)
Additional information
Additional information from genetic toxicity in vivo:
Available information on genetic toxicity:
Ames-Test:
N-(2-Nitrophenyl)phosphoric triamide induced gene mutation in Salmonella Typhimurium TA 1537 (without metabolic activation) and TA 98 (with and without metabolic activation) under the experimental conditions of the study according to OECD 471. N-(2-Nitrophenyl)phosphoric triamide is considered to be mutagenic in the bacterial reverse mutation test.
Mouse lymphoma test:
A study was performed according to OECD 476 to investigate the potential of N-(2-nitrophenyl)phosphoricacid triamide (P 101/04) to induce mutations at the mouse lymphoma thymidine kinase locus using the cell line L5178Y. The assay was performed in two independent experiments, using two parallel cultures each. During the mutagenicity test described and under the experimental conditions reported the test item did not induce mutations in the mouse lymphoma thymidine kinase locus assay using the cell line L5178Y in the absence and presence of metabolic activation. Therefore, N-(2-nitrophenyl)phosphoricacid triamide (P 101/04) is considered to be non-mutagenic in this mouse lymphoma assay.
In vivo Mouse micronucleus test:
In an in vivo mouse micronucleus test following EC-guideline B.12 under GLP conditions, the ability of N-(2-Nitrophenyl)phosphoric triamide to induce micronuclei was investigated. N-(2-nitrophenyl)phosphoric triamide did not induce micronuclei in polychromatic erythrocytes of NMRI-mice under the described experimental conditions .N-(2-nitrophenyl)phosphoric triamide is therefore considered to be non-mutagcnic in the mouse bone marrow micronucleus test.
Justification for selection of genetic toxicity endpoint
There is a positive result of an Ames test for the substance 2-NPT. However, since in an in vitro gene mutation test on mammalian cells (Mouse lymphoma assay; OECD 476) and in an in vivo mutagenicity test on somatic cells (Mouse micronucleus test; OECD 474) no indications of a mutagenic potential of the substance were found, further testing is therefore not required. Conclusion based on the following assays: Bacterial reverse mutation assay (Ames test); mouse lymphoma assay, in vivo micronucleus assay.
Justification for classification or non-classification
Based on the available data, no classification is needed.
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